PROJECT 2: TL1A, Paneth Cell & Microbial Interactions Leading to Severe Crohn's Disease Inflammatory Bowel Diseases (IBD) are the result of complex interactions between the microbial environment and susceptibility and severity genes, leading to dysregulated innate and adaptive immune responses. TNFSF15/TL1A have been shown to contribute to severity of Crohn's disease (CD). In a CD surgery cohort, we found an association between a TNFSF15 risk genotype and increased expression of TL1A in peripheral monocytes and in non-inflamed small bowel (SB) mucosa. Consistent with ?dominance? of the TL1A pathway we showed a TNFSF15 risk genotype-specific associated increase in the DR3 receptor and a decrease in TL1A antagonist DcR3 expression. We further demonstrated an association between this genotype and SB inflammation and fibrostenosis in CD, which was phenocopied in TL1A transgenic mice. Paneth cells (PC)s from patients with CD-related NOD2 variants express a lower level of antimicrobial peptides. In the above- mentioned CD surgery cohort, we found that selected NOD2 variants were also associated with alteration in granule organization and numbers with unique gene expression profiles in non-inflamed tissue. In murine models, gene variants leading to epithelial cell-specific loss of autophagy function have been associated with exacerbated SB inflammation. Variants associated with impairment of cellular stress responses were shown to result in bacteria- and/or virus-dependent PC dysfunction leading to spontaneous SB inflammation. A study showed that animals with a combination of genetic defects in both pathways developed severe mucosal inflammation. We have found significant associations between elevated TL1A and increased PC numbers/crypt. We further showed increased numbers of PCs in mice that constitutively express TL1A, with similar findings in human ileal tissues from CD patients with higher TL1A. In a stem cell generated experimental system, we showed altered PC granule phenotype in human intestinal organoids (HIOs) co-cultured with TL1A. Finally, we showed in a murine system that sustained TL1A expression required commensal microbiota to induce PC expansion. These data indicate a role for commensal microbes in TL1A driven PC changes and suggest that the intestinal microbiota, TL1A expression level, and PC function interact to modulate the spectrum of IBD severity. The evidence summarized above provides a strong rationale for studying the mechanism of TL1A- driven alteration of PCs in promoting severe SB CD and are the foundation of the hypothesis that the most severe SB CD resulting from genetically-determined overexpression of TL1A is due, at least in part, to TL1A alteration of PC development, granule structure and antimicrobial protein expression. Alterations in PC function may be the result of direct effects, or indirect effects of increased TL1A on cytokines and other molecules in conjunction with components of the microbial milieu. We further hypothesize that PC dysfunctions in NOD2 and ATG16L1 together with TL1A driven PC alteration will lead to the most severe SB CD.